In the field of regenerative medicine, research and development of three-dimensional tissues using biodegradable polymer scaffolds and cell sheets using no scaffolds are in progress. In recent years, cell sheet technologies are attracting special attention. In preparation of a cell sheet, it is necessary to put a cell suspension into cell culture vessels, grow the cells in an incubator, collect the grown monolayer cell sheets from the surfaces of the culture vessels, and stack the cell sheets. Since most cells have a nature of stopping their growth upon contacting, it is necessary to detach cell sheets from vessels and stack them in order to prepare a cell sheet having more than a certain thickness. However, protease is required to collect cells having adhesive capacity from the vessels, which leads to decomposition of the cell tissue into separate cells, and thus it has been difficult to collect cells as a normal cell tissue.
The technique by Patent Document 1 solved the above-described problem. Patent Document 1 discloses a technique in which cultured cells are collected as a cell sheet simply by using a “temperature-responsive cell culture apparatus” and changing the culture temperature. Since the cultured cells obtained have not undergone enzyme treatment, they have the advantage that they can be collected as a single sheet without destroying proteins present between cells.
However, for preparation of a cell sheet having more than a certain thickness, even the technique according to Patent Document 1 requires a laborious operation of stacking a plurality of monolayer cell sheets. There have been only a very limited number of reports in which cells were successfully formed into a three-dimensional multilayered sheet by a simple process. There has been no technique that can be used at a practical level.